Cephalometric assessment regarding craniocervical posture in orthodontic patients

A major factor that contributes to dental malocclusions is represented by the positioning of the mandible. Considering the existing interconnections between the craniocervical and craniomandibular systems it is interesting to assess how changes in one system can influence the other, thus establishing a pattern in terms of certain cephalometric landmarks that orthodontists could consider when diagnosing and evaluating an orthodontic case. Therefore, the aim of this study was to investigate the connections between cervical posture, head position, hyoid bone position in orthodontic patients with different skeletal patterns. 45 lateral cephalometric radiographs were analyzed. Skeletal class and vertical growth were the main elements that were considered when classifying patients. Craniofacial and Cervical landmarks were determined on the cephalograms, from which lines and angles resulted which were considered relevant in our study. Correlations between cephalometric variables of the patients were determined. there were some statistically significant changes identified concerning craniocervical posture and hyoid bone position between the patients in the following parameters: H-Rgn, OPT/HOR, CVT/HOR, OPT/SN, CVT/SN, H-SN. The results obtained allowed us to conclude that there were some differences at the skeletal level of the sample of patients studied. The findings are indicating that there is a close relationship between, mandible position, cervical- and head posture and the hyoid bone. The information obtained in this study could help to better understand the development of malocclusions, and to improve the orthodontic diagnostic and treatment plan.

The craniofacial complex is an extremely elaborated structure at which the forces applied during the orthodontic treatment take place. Taking into consideration that there are correlations between the craniocervical and craniomandibular systems, it is important to determine whether anatomical variations in one of these systems can cause changes in the other and vice versa, as well as to establish if these elements can be useful during orthodontic diagnosis and treatment planning 1 . Previous studies show the close relationship between the craniomandibular and craniocervical systems, both having the potential to influence each other [2][3][4][5][6] .
An anatomical element that is related both to the craniocervical system and the craniomandibular system, whose role during orthodontic treatment has not yet been fully elucidated, is the hyoid bone. The hyoid bone is a key component in the musculoskeletal system of the craniofacial complex 7 . It plays an important role in the physiological development of functions in the head and neck region such as food intake, breathing and speech, its position adapting accordingly to changes in head posture 8 . The hyoid bone is an anatomical element with an important role in the positioning of the head and neck. Previous research has shown that the hyoid bone can adapt to changes in the anteroposterior direction of the head posture and that changes of the mandible position are related with changes of the hyoid bone 9,10 . It is known that there are differences in the physiological development of functions in individuals with variations regarding their craniofacial anatomical relationships 11 . Normal functions such as mastication and breathing influence the neuromuscular system, which in turn influences the cranial posture. Thus, it was observed that the flexion and extension movements of the head that modify the Natural head position (NHP) are associated with morphological patterns 12 . Previous studies demonstrated that changes in cephalic posture related to some functional alterations such as oral respiration influences the growing patient, consequently leading to the development of typical malocclusive patterns such as mandibular retrognathia, increased gonial and craniocervical angles and causing modifications in the lingual-mandibularhyoid complex. Therefore, in oral breathing patients, cranial extensions are frequently observed. These postural changes could be a compensation for nasal airway inadequacy 13 .
As age advances, the hyoid bone may descend to lie at the level of the fourth cervical vertebra. Studies show that there is a relationship between cervical posture and dental occlusion. Patients with a class II Angle malocclusion tend to have an exaggerated kyphosis of the cervical vertebras compared to patients with normal dental occlusion 14 . This extended head posture is accompanied by a change in the resting position of the mandible and subsequently in an increase in the occlusal freeway space 15 .
A paraclinical investigation that can highlight the relationships between cervical posture and dental malocclusion is represented by the lateral cephalogram, which is a specialized radiographic technique concerned with imaging the craniofacial region in a standardized and reproductible manner 16 .
Many cephalometric analyses are available these days, but only few take into consideration the relationship established between craniocervical posture and it's connection to dental malocclusion.
Thus, aim of this study was to investigate the characteristics of craniocervical morphology in 3 groups of patients divided according to the ANB angle and according to their vertical growth pattern before starting orthodontic treatment.

Materials and methods
Study design. This was an observational and retrospective study. The sample consisted of 45 lateral cephalograms from patients aged between 25 and 30, thus considering that skeletal growth has reached maturity. Patients who have undergone any orthodontic treatment in the past were excluded from the study. Also, patients who reported systemic illnesses, musculoskeletal disorders, traumatic injuries, symptoms of TMD, complex dental restorations or symptoms of attrition were excluded. To be included in the study patients had to have lateral and anterior photographs in a natural head position (NHP) and the teleradiography had to be of good quality (also in natural head position) and to highlight at least 4 cervical vertebrae.
Ethical approval was guaranteed by the Ethics Committee at Victor Babes University of Medicine and Pharmacy Timisoara:" Aviz CECS Nr. 13/26.03.2021". All methods were performed in accordance with the relevant guidelines and regulations.
The method for obtaining the lateral cephalogram in NHP was performed as described by Meiyappan et al. 17 The lateral photographs which were also performed in an NHP, were used for superimposition, and allowed for the confirmation of a natural head position. The cephalometric software used for teleradiograph analysis with lateral photographs superimposition in NHP were performed with AudaxCeph® software (AudaxCeph Ultimate Ver 6.3.11.4346, Ljublijana, Slovenia)®.
The patients were divided into three groups, according to the ANB angle which measures the position of the maxilla in relation to the mandible in sagittal plane indicating the skeletal pattern of the patients: class I (0°-4°), class II (> 4°), class III (< 0°). In each sagittal group there were the following samples: 18 for class I; 16 for class II; 11 for class III. Also, vertical growth pattern was considered when assessing hyoid bone position. In this sense, patients were divided into 3 other categories: normodivergent (22°-28°), hyper-divergent (> 28°) and hypodivergent (< 22°), depending on the value of the FMA angle. The sample sizes for the classification regarding the vertical growth pattern were the following: 21 samples normodivergent, 14 samples hyper-divergent and 10 samples hypodivergent.
In order to evaluate the reliability of the cephalometric measurements, these were performed twice by Dr. Vlad-Tiberiu Alexa (Orthodontics and Dentofacial Orthopedics Specialist) at a one-week interval. The averages of these values found in the two measurements were used for assessment. The measurement errors were determined with respect to these averages. Fourteen days after the measurements, twelve lateral radiographs were randomly chosen for examining the error made between the two periods of tracings. The correlation coefficients between the measurement errors where greater than 0.95, demonstrating an excellent reliability. Figure 1 represents the anatomical landmarks from which lines and angles resulted. The cephalometric landmarks and their references are represented in the Table 1.

Statistical analysis.
The mean values and standard deviations of all replicates were calculated using Graph-Pad Prism (v.5.0 software, Manufacture, San Diego, CA, USA). Differences between means were analyzed with a one-way ANOVA, followed by multiple comparison analysis using the Duncan test (two-sample assuming equal variances). Differences were considered significant when p-values < 0.05. Correlations between variable was performed using Microsoft Excel 2010. Duncan tests, linear correlations between variable, principal components analysis and cluster analysis were performed using Statistica (v.12, TIBCO Software, Palo Alto, California, USA).

Results
Cephalometric differences between the groups. Hyoid bone, head position and craniocervical position in relation to certain landmarks were analyzed and the values were statistically interpreted using the Duncan test (Fig. 2). Figure 3 represents average values of the determined parameters assessing the position of the hyoid bone between patients with different vertical growth patterns. www.nature.com/scientificreports/ Correlations between variables. The analysis of correlation (Table 2) highlights a strong (r > 0.7) positive correlation between pairs I_CVT/HOR/ I_Y-axis/FK, (r = 0.737), a moderate (r > 0.5) positive correlation between the pairs I_H/ML and I_ANB (r = 0.526), I_OPT/HOR and I_H-C3 (r = 0.682). We determined mutual correlations between individual variables, which can prove dependence between parameters. The highest correlation coefficient values were determined for the I_CVT/HOR/ I_Y-axis/FK, where we can presume their mutual connection. In addition, the multi-parametric statistical evaluation method using PCA (Figs. 4 and 5), and CA ( Fig. 6) were used for further data analysis. Based on a linear correlation matrix, PCA was applied to the mean values of the measured traits to study which parameters contributed the most to total data variation.
The PCA produced 14 components. The first two principal components accounted for 23.86% and 17.05% (a total of 40.90%) of the variance, respectively. The most important variables integrated in the first component were: I_OPT/ML which was positively correlated with this component; on the other hand, I_ANB, I_Y-axis/ FK, I_CVT/HOR, I_H-SN, I_H/ML were negatively correlated with this first component. The second component was positively correlated with I_OPT/HOR and negatively correlated with I_H-ML and I_H-C3 (Fig. 4).
The similarity among class I patients was examined when each sample was plotted using the first and second principal components, which retained 40.90% of the total variance ( Fig. 4) and showed a clustering tendency, leading us to carry out correspondence analysis and depict the case through a simplified dendrogram plot. We can clearly identify class I patients with distant values by this data presentation. We were then able to put forward a dendrogram using CA, which divides class I patients into five representative clusters based on the correlation of their determined parameters.
In addition, the multi-parametric statistical evaluation method using PCA (Figs. 8 and 9), and CA ( Fig. 10) were used for further data analysis.
With the aim of studying which parameters assisted the most the total data variation, PCA was applied for the mean values of the determined traits, based on a linear correlation matrix. The PCA produced 14 components. The first two principal components accounted for 23.75% and 18.15% (a total of 41.90%) of the variance, respectively. The most important variables integrated in the first component were II_H-Rgn, II_H-C3, II_OPT/SN, II_CVT/SN, II_CVT/ML, II_OPT/ML and II_H/SN which were negatively correlated with this first component. The second component was positively correlated with II_Y-axis/FK, II_H-ML, II_H-SN and II_H/ML (Fig. 7).
The similarity among class II patients was examined when each sample was plotted using the first and second principal components, which retained 41.90% of the total variance ( Fig. 8) and showed a clustering tendency, leading us to carry out correspondence analysis and depict the case through a simplified dendrogram plot.. We www.nature.com/scientificreports/ can clearly identify class II patients with distant values by this data presentation. We were then able to put forward a dendrogram using CA, which divides class I patients into five representative clusters based on the correlation of their determined parameters.  www.nature.com/scientificreports/ Clusters classify individual patients into five groups based on similar characteristics: Group 1: II_P1, II_P15, II_P7; Group 2: II_P5, II_P8; Group 3: II_P2, II_16, II_P4; Group 4:II_P9, II_P9, II_P12, II_P14 and II_P17; and Group 5: II_P3, II_P10 and II_P11 (Fig. 9).
Regarding the parameters of III class, the analysis of correlation (Table 4)     www.nature.com/scientificreports/ In addition, the multi-parametric statistical evaluation method using PCA (Figs. 10 and 11) and CA (Fig. 12) were used for further data analysis.
Based on a linear correlation matrix, PCA was applied to the mean values of the measured traits to study which parameters contributed the most to total data variation. The PCA produced 10 components. The first two principal components accounted for 28.74% and 21.65% (a total of 50.39%) of the variance, respectively. The most important variables integrated in the first component were III_H-ML, III_H-Rgn, III_H-C3, III_OPT/SN,   (Fig. 10). The similarity among class III patients was examined when each sample was plotted using the first and second principal components, which retained 50.39% of the total variance (Fig. 11). This led us to perform CA and represent the case by a simplified dendrogram plot. Using CA, we were able to suggest a dendrogram that divides class III patients into three representative clusters based on the mutual similarity of their measured parameters (Fig. 11).

Discussion
Regarding the Y axis to Frankfurt horizontal angle (Y-axis/FK), a parameter that indicates the mandibular growth direction, significant differences between the patients from groups I-III and II-III can be observed, but no significant differences appear between I-II groups (Fig. 2). These differences can be explained by the fact that  www.nature.com/scientificreports/  13.813 mm for the class II and 16.546 mm for class III patients. There were no statistically significant differences identified, but it seems that the average distance is longer in class III patients than in class II patients which can be explained by the more anterior positioning tendency of the mandible in class III patients. The same situation was observed in the case of the angle between Hyoidale and the mandibular plane (H/ML), which  www.nature.com/scientificreports/ indicates the relation of the hyoid bone to the mandibular plane in the vertical plane. The angle between the mandibular plane and the hyoid bone is 20.333 on average for the class I skeletal patients, 21.00 for the class II and 18.818 for class III patients. As can be seen it is on average higher in class II patients and lower in class III patients which can indicate a higher position tendency of the hyoid bone in class III patients, but no significant statistically changes were identified between the 3 groups. This agrees with Amayeri et al., who did not find significant difference in the angular and linear measurements of the hyoid bone in relation to the mandible in the vertical plane 17 . The distance from H to Rgn (H-Rgn) is on average 38.889 mm for class I, 29.688 mm for class II and 41.909 mm for class III patients, showing that the distance from H to Rgn is closer for class II patients in relation with class I and further for class III patients compared with class I. This parameter indicates that there is a close connection between the hyoid bone and the positioning of the mandible in the sagittal plane in patients with different skeletal patterns. There are statistically significant differences between all skeletal classes, which can be justified by the more distal positioning of the mandible in class II patients and by the more anterior positioning of the mandible in class III patients. Also, this agrees with Amayeri et al. and Pereira Coelho et all., who found a larger distance in the position of the hyoid bone to Retrognathion in the sagittal plane for class III patients 17,18 .
The distance from H to C3 (H-C3) is on average 33.778 mm in class I, 28.688 mm in class II and 31.546 mm in class III individuals. There were statistically significant differences observed between I and II classes, but no statistical differences between pairs: I-III and II-III. However, a larger distance can be observed in class III patients than in class II patients, but which is nevertheless not statistically significant. These findings disagree with Amayeri et al. who noticed a larger distance alteration between the hyoid bone and the third cervical vertebra in Class III cases than in Class I and Class II cases, showing that there is a close link between hyoid bone-mandible-and the third cervical vertebra, which means that there is also a close connection between dental malocclusion and body posture 17 .
Cervical posture in horizontal plane differs significantly among class II and class III skeletal patients in term of inclination of the upper cervical column to the true horizontal (OPT/HOR). Values reported were: 92.889 in class I, 87.375 in class II and 95.091 in class III. This higher angle in class III patients represents the tendency of this group of patients to a more backward inclination of the cervical column while class II patients tend to have a more forward position of the column.
A significant difference can also be observed between indicators of the middle cervical segment (CVT/HOR) between the pairs I-II and II-III, but between group I-III there were no statistically significant differences. These findings indicate a mildly straighter inclination of cervical vertebra in class III and a more inclined position in class II patients. This agrees with data reported by Hedayati et al., who observed a mildly straighter inclination of the cervical vertebrae in the case of the class III patients 19 .
The average values of skeletal classes regarding the OPT/SN parameters are: 98.889 for class I, 103.500 for class II and 97.273 for class III. There are statistically significant differences in OPT/ SN, between I-II classes and II-III classes, but no differences between I-III classes. The analysis shows that the angles are wider in class II patients compared to class III patients, which means that class II patients may have a more backward head position in relation to the 2nd and 4th cervical vertebra, while class III patients have a more forward head position in relation to the 2nd and 4th cervical vertebra.
CVT/SN has the following values: 101.944 for class I, 104.500 for class II and 99.364 for class III. Significant differences can be observed between II-III classes. Also, these results highlight a more backward position of the anterior cranial base in class II patients and a more forward position of the anterior cranial base in class III patients. Hedayati et al. also reported a more forward position based on the values given by the CVT/SN angle in class III patients. Previous studies have shown that individuals with a retrognathic facial profile (class II skeletal pattern) and an obtuse cranial base angle tends to keep their heads more extended with their chins somewhat protruding, while prognathic facial profiles (class III skeletal pattern) who have a more acute cranial base angle hold their chins inclined and more inferiorly 19,20 .
The angles OPT/ML and CVT/ML have the same tendency, noticing statistically significant differences only between class I patients and class II patients. The OPT/ML angle is lower in class II patients compared to class I patients, which can be justified by a more posterior and backward positioning of the mandible in class II patients than in class I patients. However, there are some differences in the values of the OPT/ML and CVT/ML angles between class II and class III patients, but these are not statistically significant. Class III patients seem to have a higher OPT/ML and CVT/ML angles than class II patients which can be explained by a more forward positioning of the mandible. These values for the angle of the mandibular plane in relation to the cervical column could be explained by the fact that mandibular growth is different among individuals and the mandibular rest position, besides cervical and head posture is also dependent on several factors such as occlusal interferences, temporomandibular dysfunction, psychosocial stress, diurnal variation, and nasal obstruction [21][22][23][24] . Also, the counterclockwise rotation of the mandible which is more prevalent in class II patients and the anticlockwise rotation of the mandible, which is more prevalent in class III patients, plays a significant role in determining these angles.
Regarding the linear distance between the anterior cranial base and the hyoid bone (H-SN) there were statistically differences between classes II and III. The values are: 93.611 for class I, 94.00 for class II and 104.546 for class III. The analysis shows that there is a greater H-SN distance in class II patients compared to class I and even greater distance in class III patients compared to class I patients. This means that the hyoid bone position is lower in the vertical plane in relation to the anterior cranial base in class III cases than in class II and I case. This agrees with Amayeri  www.nature.com/scientificreports/ posterior position of the hyoid bone in Class II skeletal malocclusion subjects and with Adamidis and Spyropoulos, who reported a more anterior position of the Hyoid bone in class III patients [25][26][27] . This can be attributed to the muscular attachment of the hyoid bone and the mandible. The hyoid follows the mandibular movements in the sagittal plane, so it moves backwards in class II individuals and forward in class III patients. Natural head posture is the upright position of the head of an individual while the head is in a balanced by the post-cervical, suprahyoid and infrahyoid muscle group 28 . It is dependent on the respiratory function, visual function, and the masticatory muscles. Our cephalometric findings revealed some relationships between the skeletal pattern of a patient, the hyoid bone, head posture and the cervical vertebras. Respectively, we demonstrated that class II patients tend to have a more accentuated cervical kyphosis, with their head position inclined more backward, while class III patients have a more accentuated cervical lordosis with their head position inclined more forward. These findings could explain the relationship between the cervical column posture, head posture and orthodontic malocclusions. The backward positioning of the head in class II patients, may be because of the patient's attempt to compensate the exaggerated kyphosis of the cervical spine resulting in a more distally positioning of the mandible, while in class III patients the more forward positioning of the head, could be the result of the patient's attempt to compensate for the exaggerated cervical lordosis resulting in a more anteriorly positioning of the mandible.
Regarding the position of the hyoid bone between patients with different vertical growth patterns respectively normodivergent, hyperdivergent and hypodivergent (Fig. 3) according to the FMA angle, our research has revealed some significant differences in some of the parameters evaluated. Vertical position of the hyoid bone in individuals with hyperdivergent growth pattern was slightly lower compared to hypodivergent and normodivergent patients. H-SN value was 93.95 for normodivergent patients, 104.78 for hyperdivergent patients and 89.40 for hypodivergent patients. There were statistically significant differences found in between normodivergent patients and hyperdivergent patients but also between hyperdivergent patients and hypodivergent patients. This agrees with Bhullar et al. who also found that the vertical position of the hyoid bone is slightly uppward in hypodivergent growth pattern patients compared to hyperdivergent patients 29 . The

Conclusions
Consequently, the most important findings of our paper are: • Hyoid bone position in relation to retrognathion is closer in class II patients than in class III patients.
• Cervical position differs significantly between class II and class III patients in terms of cervical spine inclination. • Class III patients have a cervical lordosis tendency, while in class II patients a cervical kyphosis tendency can be observed. • Class II patients have a more backward inclination of the head, this having a detrimental effect at the occlusal level, through a more posterior positioning of the mandible. • In class III patients there is an opposite effect. The position of the head being more down and forward most likely as a way of compensating the body for tilting the cervical spine further backwards resulting in a more anterior position of the mandible.
These findings indicate a close relationship between, hyoid bone positioning, cervical and head posture. Therefore, affected patients may compensate for pathological changes at the skeletal level through postural adjustments of the head and neck region. To further evaluate possible applications for orthodontic diagnosis and treatment, more studies should be carried out in this field and to evaluate their possible connections with dental malocclusions. www.nature.com/scientificreports/ Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http:// creat iveco mmons. org/ licen ses/ by/4. 0/.